It is well known in the art that it is possible to obtain a cell line which is able to produce a homogenous, i.e. monoclonal, antibody. The basic technique (Kohler and Milstein, Nature, 256, 1975) comprises the fusion of mouse myeloma cells with spleen cells to form hybridoma cells and selection from these of clones capable of producing the desired antibody. This general procedure has also been described in U.S. Pat. Nos. 4,364,932, 4,364,934, 4,364,935, 4,364,937 and 4,361,550.
Although the general method has been known for some years, the preparation and selection of each suitable hybridoma presents its own special difficulties. There is indeed no certainty that a suitable hybridoma will be found and, equally, there is no certainty that the hybridoma will produce an antibody having the desired properties.
Monoclonal antibodies have a variety of uses, in particular for the isolation and purification of the proteins to which they are specific or for assaying them e.g. in a diagnostic kit; see for example PCT published applications W081/02899 and W082/01773.
Whereas the family of human .alpha.-interferons is coded for by a number of distinct genes and thus shows variations in the amino acid sequences, human .gamma.-interferon is coded for by only a single gene. However, human .gamma.-interferon is a very labile protein and degrades in particular through shortening of its carboxy terminus to yield a number of smaller .gamma.-interferons. Thus the complete amino acid sequence of human .gamma.-interferon (GIF), as deduced from its cDNA sequence, is:
__________________________________________________________________________ S1 S10 met lys tyr thr ser tyr ile leu ala phe gln leu cys ile val S20 1 10 leu gly ser leu gly CYS TYR CYS GLN ASP PRO TYR VAL LYS GLU 20 ALA GLU ASN LEU LYS LYS TYR PHE ASN ALA GLY HIS SER ASP VAL 30 40 ALA ASP ASN GLY THR LEU PHE LEU GLY ILE LEU LYS ASN TRP LYS 50 GLU GLU SER ASP ARG LYS ILE MET GLN SER GLN ILE VAL SER PHE 60 70 TYR PHE LYS LEU PHE LYS ASN PHE LYS ASP ASP GLN SER ILE GLN 80 LYS SER VAL GLU THR ILE LYS GLU ASP MET ASN VAL LYS PHE PHE 90 100 ASN SER ASN LYS LYS LYS ARG ASP ASP PHE GLU LYS LEU THR ASN 110 TYR SER VAL THR ASP LEU ASN VAL GLN ARG LYS ALA ILE HIS GLU 120 130 LEU ILE GLN VAL MET ALA GLU LEU SER PRO ALA ALA LYS THR GLY 140 LYS ARG LYS ARG SER GLN MET LEU PHE ARG GLY ARG ARG ALA SER 146 GLN. __________________________________________________________________________
The leader sequence consists of the 20 amino acid residues shown in small print and probably of the next three also. The various .gamma.-interferons identified herein are given in the following Table:
______________________________________ Interferon Type Amino Acids ______________________________________ .gamma.-interferon A or GIFA 1-146 .gamma.-interferon A' or GIFA' 1-142 .gamma.-interferon B or GIFB* 1-131 .gamma.-interferon D or GIFD 4-146 ______________________________________ *GIFB is relatively inactive.
GIFA can be cleaved by CNBr at the methionine residues (amino acids 48, 80, 120 and 137). The resulting mixture of five cleavage products will be referred to as "CNBr-GIFA" herein.
In addition, a peptide consisting of the fifteen carboxy-terminal amino acids, 132-146, and identified herein as 15AA peptide, was used in the test procedures described herein.
The .gamma.-interferons (GIFs) have in various tests proved to be most promising pharmaceutically active substances useful for combatting various diseases, in particular viral diseases and certain forms of cancer. .gamma.-Interferon is widely described in the literature and may be obtained either by mitogenic induction of lymphocytes or by recombinant DNA technology, e.g. as described in the published European Patent Applications Nos. 77,760, 88,540 and 95,350.